The goal of the research proposed in this application is control of the endogenous mouse genome by elements derived from the lac operon of E. coli. It is a direct extension of the goal of the applicants' original proposal, which was to gain control of murine transgenes using these same elements. Even as they complete this goal, the applicants are aware that the ultimate target of their efforts must be genes in their natural environment, where all the factors that regulate and impinge on activity are present. Although the appicants' personal reason for developing this system is to understand the role of gene regulation in development and plasticity, the system will be a powerful tool for understanding the molecular underpinning of a wide range of biological and biomedical phenomena. The focus of the original grant was the development of a mouse that expresses functional levels of the lac repressor, the key regulatory protein of the murine lac system. The focus of this continuation is on the second component of the system, the unique promoter element recognized by the repressor, the lac operator. Two copies of this 26 bp sequence approximately 200 bp apart can confer the ability to regulate promoters of mammalian genes (p53) or mammalian promoters driving cDNA (tyrosinase) when introduced as transgenes into the murine genome. The next obvious step is to introduce these lac operators into the promoter of an endogenous gene by homologous recombination (Specific Aim #1). Targeted alleles could be controlled globally, by crossing onto a background in which the lac repressor is ubiquitously expressed, such as the one that the applicants have already developed, or in a cell- or tissue-specific manner using lac repressors with restricted expression patterns (Specific Aim #2). By the end of the second phase of the project, the goal is to have transferred the lac system from the transgenic mouse to the endogenous genome and to have developed a relatively simply methodology by which even complex phenotypes can be modeled (Specific Aim #3).